J Antimicrob Chemother 2020; 75: 300–308 doi:10.1093/jac/dkz434 Advance Access publication 21 October 2019 Mutations that increase expression of the EmrAB-TolC efflux pump confer increased resistance to nitroxoline in Escherichia coli Downloaded from https://academic.oup.com/jac/article-abstract/75/2/300/5601609 by Uppsala Universitetsbibliotek user on 01 April 2020 Fabiola Pue´rtolas-Balint1, Omar Warsi1, Marius Linkevicius1,2, Po-Cheng Tang 1,3 and Dan I. Andersson1* 1Department of Medical Biochemistry and Microbiology, Uppsala University, SE-75123, Uppsala, Sweden; 2Department of Public Health Solutions, National Institute for Health and Welfare (THL), Helsinki, Finland; 3Department of Medical Cell Biology, Uppsala University, SE-75123, Uppsala, Sweden *Corresponding author. E-mail:
[email protected] Received 10 June 2019; returned 17 July 2019; revised 2 September 2019; accepted 19 September 2019 Objectives: To determine the mechanism of resistance to the antibiotic nitroxoline in Escherichia coli. Methods: Spontaneous nitroxoline-resistant mutants were selected at different concentrations of nitroxoline. WGS and strain reconstruction were used to define the genetic basis for the resistance. The mechanistic basis of resistance was determined by quantitative PCR (qPCR) and by overexpression of target genes. Fitness costs of the resistance mutations and cross-resistance to other antibiotics were also determined. Results: Mutations in the transcriptional repressor emrR conferred low-level resistance to nitroxoline [nitroxoline MIC (MICNOX)=16 mg/L] by increasing the expression of the emrA and emrB genes of the EmrAB-TolC efflux pump. These resistant mutants showed no fitness reduction and displayed cross-resistance to nalidixic acid. Second-step mutants with higher-level resistance (MICNOX=32–64 mg/L) had mutations in the emrR gene, to- gether with either a 50 kb amplification, a mutation in the gene marA,oranISupstreamofthelon gene.